Journal of Leukocyte Biology 43:80-90 (1988) © 1988 Alan R. Liss, Inc. Pulmonary Interstitial Macrophages: Isolation and Flow Cytometric Comparisons With

Pulmonary interstitial macrophages (iM) were isolated from rat lungs by an Fc-y receptor-based affinity technique coupled with multiparameter flow cytometry. Single cell sus-pensions obtained by collagenase digestion of extensively perfused and lavaged lungs were applied to carpets of opsonized sheep red blood cells (SRBC-lgG) bound to plastic tissue culture flasks. At 0-4#{176}C,optimal binding of lung cells occurred within 60 mm at plating densities of i-2 x 106 lung cells/cm2 when the SRBC substrate was opsonized with iO g/ml anti-SRBC lgG. Nonadherent cells were removed by gently rinsing the plates and adherent cells were recovered by lysing the SRBC-lgG substrata. By light microscopy, the mixture of adherent cells was comprised of mononuclear cells (-54%), many of which appeared to be macrophages, lymphocytes (- 20%), polymorphonuclear leukocytes (- 15%), plasma cells (- 8%), eosinophils (- 2%), and mast cells (- 0.5%). The cells which adhered to the SRBC-lgG monolayers were further resolved into subpop-ulations by multiparameter flow cytometry and sorted according to their electro-optical characteristics. One subpopulation appeared morphologically to be macrophages, and> 90 % of these cells readily phagocytized SRBC-lgG in vitro. Peroxidase staining of this population was minimal, indicating that these cells were not blood monocytes (BM)